The goal of the studies described in this proposal is to characterize the recognition site in brain for methylphenidate (a centrally acting stimulant) and to investigate its relationship to the dopamine transport complex. A newly synthesized tritiated form of methylphenidate will be used in this research. Initial studies with this radioligand (Schweri, et al. and Janowsky et al., Appendix I) suggest that the binding site for methylphenidate may be associated with the dopamine transport system. The present studies are designed to further explore this possibility using a number of approaches. Biochemical studies will determine the regional distribution and ionic modulation of methylphenidate binding, involvement of critical amino acid residues in the binding site, and nature of the inhibition of binding by dopamine. Pharmacological studies will correlate inhibition of methylphenidate binding by specific drug classes with potency to block dopamine uptake in order to pinpoint the biological significance of this binding site. Especially exciting are plans to determine if an analogue of phencyclidine which has been found to irreversibly inhibit methylphenidate binding will block dopamine transport in the same manner. The physiologic relevance of this binding site will be established by investigating age, sex, strain and species differences in methylphenidate binding and dopamine uptake. Other experiments will explore homeostatic regulation of methylphenidate binding in response to alterations in the synaptic concentration of dopamine. Finally, methylphenidate binding will be studied in an animal model of hyperactivity. The results of this research will further our understanding of the relationship between the recognition site for stimulants and the dopamine transport complex. In the short term this may facilitate studies in post mortem tissues where actual measurement of dopamine uptake is not feasible. Ultimately, it may afford new ways to manipulate the activity of dopaminergic pathways. This holds important implications for the treatment of dopamine-related disorders such as schizophrenia, narcolepsy, Parkinsons's disease, and hyperactivity.